RESUMO
The relation of ethambutol resistance to embB mutations remains unclear, and there are no reports on ethambutol resistance from the caribbean. We examined the sequence of embB in 57 distinct Multi-Drug Resistant (MDR) and non-MDR strains of Mycobacterium tuberculosis, mostly from Cuba and the Dominican Republic. embB306 codon mutations were found exclusively in MDR-TB, but in both ethambutol sensitive and resistant strains. Valine substitutions predominated in ethambutol resistant strains, while isoleucine replacements were more common in sensitive strains. Three ethambutol resistant MDR strains without embB306 substitutions had replacements in embB406 or embB497, but these were also found in ethambutol sensitive MDR strains. The results confirm previous findings that amino acid substitutions in EmbB306, EmbB406 and EmbB497 are found only in MDR-TB strains but in both phenotypically resistant and sensitive strains. One ethambutol resistant non-MDR strain did not have any embB mutation suggesting that other undefined mutations can also confer ethambutol resistance.
Assuntos
Antituberculosos/farmacologia , Resistência Microbiana a Medicamentos/genética , Etambutol/farmacologia , Mycobacterium tuberculosis/genética , Pentosiltransferases/genética , Tuberculose Resistente a Múltiplos Medicamentos/microbiologia , Substituição de Aminoácidos , Códon/genética , Cuba/epidemiologia , Análise Mutacional de DNA , DNA Bacteriano/genética , República Dominicana/epidemiologia , Relação Dose-Resposta a Droga , Farmacorresistência Bacteriana Múltipla/genética , Humanos , Testes de Sensibilidade Microbiana , Mutação , Mycobacterium tuberculosis/efeitos dos fármacos , Pentosiltransferases/fisiologia , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Tuberculose Resistente a Múltiplos Medicamentos/epidemiologiaRESUMO
The relation of ethambutol resistance to embB mutations remains unclear, and there are no reports on ethambutol resistance from the caribbean. We examined the sequence of embB in 57 distinct Multi-Drug Resistant (MDR) and non-MDR strains of Mycobacterium tuberculosis, mostly from Cuba and the Dominican Republic. embB306 codon mutations were found exclusively in MDR-TB, but in both ethambutol sensitive and resistant strains. Valine substitutions predominated in ethambutol resistant strains, while isoleucine replacements were more common in sensitive strains. Three ethambutol resistant MDR strains without embB306 substitutions had replacements in embB406 or embB497, but these were also found in ethambutol sensitive MDR strains. The results confirm previous findings that amino acid substitutions in EmbB306, EmbB406 and EmbB497 are found only in MDR-TB strains but in both phenotypically resistant and sensitive strains. One ethambutol resistant non-MDR strain did not have any embB mutation suggesting that other undefined mutations can also confer ethambutol resistance.
Assuntos
Antituberculosos/farmacologia , Etambutol/farmacologia , Mycobacterium tuberculosis/genética , Pentosiltransferases/genética , Resistência Microbiana a Medicamentos/genética , Tuberculose Resistente a Múltiplos Medicamentos/microbiologia , Análise Mutacional de DNA , Cuba/epidemiologia , Códon/genética , DNA Bacteriano/genética , Farmacorresistência Bacteriana Múltipla/genética , Humanos , Mutação , Mycobacterium tuberculosis/efeitos dos fármacos , Pentosiltransferases/fisiologia , Relação Dose-Resposta a Droga , Reprodutibilidade dos Testes , República Dominicana/epidemiologia , Sensibilidade e Especificidade , Substituição de Aminoácidos , Testes de Sensibilidade Microbiana , Tuberculose Resistente a Múltiplos Medicamentos/epidemiologiaRESUMO
The direct detection of pyrazinamide resistance in Mycobacterium tuberculosis is sufficiently difficult that many laboratories do not attempt it. Most pyrazinamide resistance is caused by mutations that inactivate the pyrazinamidase enzyme needed to convert the prodrug pyrazinamide to its active form. We evaluated two newer and simpler methods to assess pyrazinamidase activity, the nitrate reductase and malachite green microtube assays, using nicotinamide in place of pyrazinamide. A total of 102 strains were tested by these methods and the results compared with those obtained by the classic Wayne assay. Mutations in the pncA gene were identified by sequencing the pncA genes from all isolates in which pyrazinamide resistance was detected by any of the three methods. Both the nitrate reductase and malachite green microtube assays showed sensitivities of 93.75% and specificities of 97.67%. Mutations in the pncA gene were found in 14 of 16 strains that were pyrazinamide resistant and in 1 of 4 strains that were sensitive by the Wayne assay. Both of these simple methods, used with nicotinamide, are promising and inexpensive alternatives for the rapid detection of pyrazinamide resistance in limited-resource countries.
Assuntos
Antituberculosos/farmacologia , Colorimetria/métodos , Farmacorresistência Bacteriana , Mycobacterium tuberculosis/efeitos dos fármacos , Niacinamida/metabolismo , Pirazinamida/farmacologia , Amidoidrolases/genética , Humanos , Testes de Sensibilidade Microbiana/métodos , Nitratos/metabolismo , Compostos Organometálicos/metabolismo , Sensibilidade e Especificidade , Análise de Sequência de DNARESUMO
BACKGROUND: Conventional methods for susceptibility testing require several months before results can be reported. However, rapid methods to determine drug susceptibility have been developed recently. Phage assay have been reported as a rapid useful tools for antimicrobial susceptibility testing. The aim of this study was to apply the Phage assay for rapid detection of resistance on Mycobacterium tuberculosis strains in Cuba. METHODS: Phage D29 assay was performed on 102 M. tuberculosis strains to detect rifampicin resistance. The results were compared with the proportion method (gold standard) to evaluate the sensitivity and specificity of Phage assay. RESULTS: Phage assay results were available in 2 days whereas Proportion Methods results were obtain in 42 days. A total of 44 strains were detected as rifampicin resistant by both methods. However, one strains deemed resistant by Proportion Methods was susceptible by Phage assay. The sensitivity and specificity of Phage assay were 97.8 % and 100% respectively. CONCLUSION: Phage assay provides rapid and reliable results for susceptibility testing; it's easy to perform, requires no specialized equipment and is applicable to drug susceptibility testing in low income countries where tuberculosis is a major public health problem.